Tandem Mass Spectrometry-Based Amyloid Typing Using Manual Microdissection and Open-Source Data Processing

Objectives Standard implementations of amyloid typing by liquid chromatography-tandem mass spectrometry use capabilities unavailable to most clinical laboratories. To improve accessibility of this testing, we explored easier approaches to tissue sampling and data processing. Methods We validated a typing method using manual sampling in place of laser microdissection, pairing the technique with a semiquantitative measure of sampling adequacy. In addition, we created an open-source data processing workflow (Crux Pipeline) for clinical users. Results Cases of amyloidosis spanning the major types were distinguishable with 100% specificity using measurements of individual amyloidogenic proteins or in combination with the ratio of lambda and kappa constant regions. Crux Pipeline allowed for rapid, batched data processing, integrating the steps of peptide identification, statistical confidence estimation, and label-free protein quantification. Conclusions Accurate mass spectrometry-based amyloid typing is possible without laser microdissection. To facilitate entry into solid tissue proteomics, newcomers can leverage manual sampling approaches in combination with Crux Pipeline and related tools.